Rapid developments in the field of recombinant DNA technology in recent years have made it possible to produce a wide range of biologically important polypeptides and proteins in high yields. Essential to such production is the availability of DNA sequences coding for the desired products. Generally, such DNA sequences are produced enzymatically using populations of messenger RNA (mRNA) isolated from cells known to produce the desired polypeptide or protein as templates. Complementary DNA strands are first produced from the mRNA templates, and second DNA strands are then produced using the first DNA strands as templates.
The result is a mixture of duplex complementary DNA (cDNA) molecules which together comprise a cDNA "library". The cDNAs in such libraries can be replicated through cloning in appropriate host cells and identified by the use of molecular probes, antibodies or biological activity assays.
A number of scientific investigators have devised methods for producing cDNA libraries. One widely used method is that of Okayama et al. [Mol. Cell. Biol. 2:161 (1982)]. This method employs an oligo (dT)-tailed plasmid primer and an oligo (dG)-tailed linker DNA. Applying the method to rabbit reticulocyte mRNA, Okayama et al. produced about 10.sup.5 globin cDNA clones per .mu.g of mRNA. More recently, using highly competent cells, the Okayama et al. method has been found to yield 1 to 2.times.10.sup.6 clones per .mu.g of plasmid vector [Okayama et al., Meth. Enzymol. 154:3 (1987); Margolskee et al., Mol. Cell. Biol. 8:2837 (1988)]. Gubler et al. [Gene 25:263 (1983)] reported a method for generating cDNA libraries which avoids the use of plasmid primers and produces a cloning efficiency of as high as 10.sup.6 recombinants per .mu.g of mRNA. These and other cDNA cloning methods have been reviewed by Kimmel et al. [Meth. Enzymol. 152:307 (1987)].
Although the foregoing methods have been used to make cDNAs coding for a variety of polypeptides and proteins, the efficiency of such methods may be inadequate if a desired mRNA species is present in relatively low abundance in the total isolated mRNA population. For the cloning of such minor species, substantially higher cloning efficiencies are required.